The transformation of cholesterol to pregnenolone by adrenal mitochondrial preparations will be studied qualitatively and quantitatively. We will, specifically, prepare and test 20(22)-dehydrocholesterol, 20(22)-oxidocholesterol and (20S)-20-hydroperoxycholesterol as precursors of pregnenolone. We will investigate proposed kinetic data, particularly in view of a suggested compartmental division of enzyme and substrate which will be pertinent for the study of the action of trophic hormones. The quantitative aspects of compartmentalization will be studied with molecules marked with deuterium and tritium, while kinetic data will be obtained with sterols marked with tritium and carbon-14. Cholesterol analogs with a variety of substituents such as methyl, ethyl, vinyl, amino and fluoro at C-20 and C-22 will be produced and their biological activity tested for inhibition of cholesterol and corticoid production.